1. Field of the Invention
The present invention relates to a mass spectrometry method and a mass spectrometry device using an ion trap mass spectrometer.
2. Description of the Related Art
A mass spectrometer is an instrument that ionizes a sample molecule by applying an electric charge thereto and separates the generated ions according to their mass to charge ratio by an electric field or by a magnetic field, and then measures the quantity as a current value with a detector.
A mass spectrometer has high sensitivity and is also excellent in quantitative and identification capabilities as compared to the conventional analyzers. In recent years, peptide analysis carried out in place of genomic analysis attracts an attention in the field of life science, and thus effectiveness of a mass spectrometer, which has high sensitivity and is excellent in identification capability, has been reassessed.
The measurement of a sample by a mass spectrometer gives a current value in the unit of mass to charge ratio. This is referred to as a mass spectrum. This mass spectrum varies according to the structure of a sample to be measured, and information on the structure of the sample can be obtained from the pattern of this mass spectrum.
However, in some cases, constituents within a sample are complicated or the obtained mass spectrum is information insufficient for identifying the constituents.
Especially in a mass spectrometer, molecular ions are separated according to their mass to charge ratios, so if the mass to charge ratios are the same in spite of different structures, it is difficult to distinguish the molecular ions. In order to solve this, an MSn analysis is devised.
In the MSn analysis: molecular ions are introduced into a mass spectrometer; ions other than an ion having a specific mass number are removed from the mass spectrometer; a collision between the selected molecular ion thereby and a neutral molecule is caused to break the bond of a part of the molecular ion; and the ions having the broken bond are measured.
The breaking of the bond of a selected molecular ion, which is caused to collide with this neutral molecule, is referred to as collision induced dissociation (CID), and is referred to as MS2, MS3 and the like according to the number of repetitions of a series of operations of ion accumulation, ion isolation and collision induced dissociation.
For the bond between atoms within a molecule, the bond energy varies according to the structure thereof and the type of bond, and therefore a portion having a lower bond energy is likely to be broken by collision induced dissociation.
During a collision between a molecular ion and a neutral molecule, a kinetic energy required for breaking the bond is applied to the molecular ion to thereby form fragment ions peculiar to the molecular ion, allowing the structure of the molecular ion to be known.
Moreover, as the number of repetitions of MSn analysis increases, more structure information can be obtained.
As a mass spectrometer, various types are listed according to the operation and configuration for separating ions, and one of the configurations suitable for conducting MSn analysis is an ion trap mass spectrometer.
The ion trap mass spectrometer can carry out ion isolation and collision induced dissociation by forming such quadrupolar electric field that ions having a particular mass to charge ratio will stay in an ion trap, and then by varying the quadrupolar electric field.
When a first round of collision induced dissociation is carried out, the resultant ions produced may be subjected to a second round of ion isolation and of collision induced dissociation instead of being induced to a detector. In this way, it is possible to conduct a plurality of MSn analyses.
The operation of ion trap is mainly divided into four operations in conducting the MSn analysis.
1. Apply a main high frequency voltage to a ring electrode to form a quadrupolar electric field in the ion trap.
Since ions generated outside or inside of the ion trap have an electric charge, if the ions are introduced in the quadrupolar electric field, the ions with a mass to charge ratio corresponding to a value and q value that exist in the region of a hatched portion in a stability region diagram shown in FIG. 1 will draw a stable trajectory and be accumulated in the ion trap.
These a value and q value are determined by Equation (1) using the main high frequency voltage V, the main direct current voltage U, the inner radius r0 of the ring electrode, the angular frequency Ω of the main high frequency voltage, and the mass to charge ratio m/z of the ion.
                                          q            z                    =                                    4              ⁢                                                          ⁢              z              ⁢                                                          ⁢              V                                      (                              m                ⁢                                                                  ⁢                                  r                  0                  2                                ⁢                                  Ω                  2                                            )                                      ⁢                                  ⁢                              α            z                    =                                    8              ⁢                                                          ⁢              z              ⁢                                                          ⁢              U                                      (                              m                ⁢                                                                  ⁢                                  r                  0                  2                                ⁢                                  Ω                  2                                            )                                                          Equation        ⁢                                  ⁢                  (          1          )                    
The operation to accumulate ions in the ion trap in this manner is referred to as the ion accumulation operation.
Since in the ion accumulation operation, the operation is carried out under a condition of a=0 without applying the main direct current voltage (U), ions to be accumulated are uniquely determined on a=0 line in the stability region diagram according to their mass to charge ratio.
The q value is in the range of 0 to 0.908, and the ions having the mass to charge ratio corresponding to this range are accumulated in the ion trap.
2. The accumulated ions draw a stable trajectory at respective natural frequencies of plural components according to their mass to charge ratio in the ion trap. ω0, which is one of the frequency components, can be estimated from a βz value shown in the stability region diagram of FIG. 1 and Equation (2).
                              ω          0                =                                            β              z                        ⁢            Ω                    2                                    Equation        ⁢                                  ⁢                  (          2          )                    
If an auxiliary alternate current voltage corresponding to this frequency is applied to an end cap electrode, ions will resonate and be ejected from the ion trap by an auxiliary alternate current electric field generated in the ion trap.
If this operation is carried out to the undesired ions not subjected to structure analysis, i.e., not subjected to collision induced dissociation, the ions will be ejected from the ion trap and only the desired ions will be accumulated in the ion trap. This is referred to as the ion isolation operation.
3. Then, if a frequency, which resonates with the desired ions subjected to collision induced dissociation, is applied to the end cap electrode to such an extent that the desired ions are not ejected, then the ions obtain a potential by the auxiliary alternate current electric field and repeats a collision with the neutral molecule inside the ion trap, so that a bond in the ion is broken to generate fragment ions. This is referred to as the collision induced dissociation operation.
4. The repeating of these ion isolation operation and collision induced dissociation operation allows for MSn analysis, and by operating the ring electrode and the end cap electrode after the fragment ions worth obtaining the structure information are accumulated in the ion trap, the ions reach a detector according to their mass to charge ratio, where the ion quantity corresponding to the mass to charge ratio is detected as a current value.
The examples indicating such ion trap operation include Japanese Patent Translation Publication. No. 9-501536, Japanese Patent Application Laid-Open Publication No. 2002-184348, Japanese Patent Application Laid-Open Publication No. 2002-313276 and the like.